MOLECULAR MECHANISMS OF GENETIC RECOMBINATION

本文提出的研究的长期目标是确定 同源重组的分子机制，通过研究酶， 促进该进程的各个步骤和 每一步的DNA中间体。 另一个目标是确定 调节重组的机制。 这项研究将 专注于大肠杆菌中重组的RecBCD途径， 粟酒裂殖酵母减数分裂重组 研究的具体目标是：大肠杆菌是为了1）验证一个小RNA 一个子单元，似乎需要一些，但不是所有的活动， RecBCD酶，2）测定RecBCD酶的改变， 切气，导致失去一些但不是所有的活动， 3)测试RecBCD酶的DNA解旋模型，基于 RecBCD酶-DNA起始复合物的结构，以及4）分析 在RecBCD中形成的含杂合DNA的中间体的结构 通路 这些目标将通过基因和 酶学研究 研究的具体目标是在S。粟酒裂殖子是1）分析减数分裂 重组缺陷（rec）突变体以确定它们的主要缺陷， 2)通过克隆测定rec基因产物的活性， 对基因进行测序，纯化它们的产物， 重组可能需要的活动，以及3）测试角色 酶，如核酸外切酶I和II，可能需要 通过产生缺乏这些酶的突变体进行重组。 生物多样性是生物多样性的重要组成部分， 有机体和细胞水平。 重组的异常产生 染色体重排，如缺失和易位。 染色体重排与以下疾病有关，也可能是其原因之一， 出生缺陷和癌症。 了解的分子机制 基因重组对于确定这些疾病的病因很重要 并有可能阻止他们。

The long-term objective of the research proposed here is to determine the molecular mechanisms of homologous recombination, by studying the enzymes promoting the individual steps of the process and the structures of the DNA intermediates at each step. An additional objective is to determine the mechanisms by which recombination is regulated. The research will focus on the RecBCD pathway of recombination in Escherichia coli and on meiotic recombination in Schizosaccharomyces pombe. Specific aims of the research in E. coli are to 1) verify a small RNA subunit that appears to be required for some but not all activities of the RecBCD enzyme, 2) determine the alteration in RecBCD enzyme upon cutting at Chi, resulting in loss of some but not all of its activities, 3) test a model of DNA unwinding by RecBCD enzyme, based upon the structure of the RecBCD enzyme-DNA initiation complex, and 4) analyze the structure of hybrid DNA-containing intermediates formed in the RecBCD pathway. These goals will be sought by a combination of genetic and enzymatic studies. Specific aims of the research in S. pombe are to 1) analyze meiotic recombination-deficient (rec) mutants to determine their primary defects, 2) determine the activities of the rec gene products, by cloning and sequencing the genes, purifying their products, and testing them for activities likely to be required for recombination, and 3) test the roles of enzymes, such as exonucleases I and II, likely required for recombination by generating mutants lacking these enzymes. Recombination is important for generating diversity at both the organismal and cellular levels. Aberrancies of recombination generate chromosomal rearrangements, such as deletions and translocations. Chromosomal rearrangements are associated with, and may be a cause of, birth defects and cancers. Understanding the molecular mechanisms of recombination is important in determining the causes of these diseases and possibly preventing them.